Monoclonal antibodies are an extremely valuable tool for studying proteins. We will use genetic, biochemical, and physical methods to define how a panel of monoclonal antibodies bind to the N-terminal domain of the phage lambda repressor. The three-dimensional structure of the N-terminal domain is known and it has been the subject of intensive genetic and biochemical study. We will use mutant repressors with surface substitutions to determine which regions of the protein are involved in antibody recognition. Some of these mutant repressors currently exist; others will be created by oligonucleotide-directed mutagenesis; and some will be selected directly by screening with the antibodies. In some cases, we will attempt to co- crystallized the antibody FAB fragment and the N-terminal domain. Interactions between the antibodies and synthetic peptides will also be investigated to determine the conformational stability of these peptides. We will use the monoclonal antibodies as reagents for detecting amino acid substitutions that restore the ability of unstable mutant repressors to fold into a stable, protease resistant structure. In an otherwise wild-type background, many of these reverting substitutions should increase the stability of the N- terminal domain to thermal denaturation or denaturation by urea or guanidine-HCL.